1. Field of the Invention
The present invention relates to methods of producing secreted polypeptides having biological activity and to fusion proteins and polynucleotides thereof.
2. Description of the Related Art
The recombinant production of a heterologous polypeptide in a fungal host cell, particularly a filamentous fungal cell such as Aspergillus or Trichoderma or a yeast cell such Saccharomyces, may provide for a more desirable vehicle for producing the polypeptide in commercially relevant quantities.
Recombinant production of a secreted heterologous polypeptide is generally accomplished by constructing an expression cassette in which the DNA coding for the polypeptide is operably linked to a promoter suitable for the host cell and a signal peptide coding region that codes for an amino acid sequence linked in frame to the amino terminus of the polypeptide and directs the encoded polypeptide into the cell's secretory pathway. The expression cassette is introduced into the host cell, usually by plasmid-mediated transformation. Production of the secreted heterologous protein is then achieved by culturing the transformed host cell under inducing conditions necessary for the proper functioning of the promoter contained on the expression cassette.
While expression of a heterologous polypeptide in a host cell may be improved, an obstacle often encountered is that the polypeptide is poorly secreted into the culture medium. One method of improving secretion of the polypeptide is to replace the native signal peptide coding sequence with a foreign signal peptide coding region to enhance secretion of the polypeptide. However, in some cases, such a replacement does not provide a sufficient improvement for producing the polypeptide in commercially relevant quantities. Another method is to fuse the polypeptide to another polypeptide that is highly secreted by a host cell. The highly secreted polypeptide functions as a carrier to transport the poorly secreted or non-secreted polypeptide as a fusion protein through the cell's secretory pathway.
WO 05/093050 discloses a fusion protein composed of an exo-cellobiohydrolase catalytic domain and a cellulase catalytic domain to increase the yield of a cellulase enzyme. Gouka et al., 1997, Applied and Environmental Microbiology February 1997, p. 488-497, discloses glucoamylase gene fusions that alleviate limitations for protein production in Aspergillus awamori. Nyyssonen and Keranen, 1995, Current Genetics 28; 71-79, discloses multiple rotes of the cellobiohydrolase I in enhancing production of fusion antibodies by Trichoderma reesei. 
It is an object of the present invention to provide methods for increasing the secretion of polypeptides having biological activity.